Stereoselective Olefin Cyclopropanation under Aerobic Conditions with an Artificial Enzyme Incorporating an Iron-Chlorin e6 Cofactor

Myoglobin has recently emerged as a promising biocatalyst for catalyzing carbene-mediated cyclopropanation, a synthetically valuable transformation not found in nature. Having naturally evolved for binding dioxygen, the carbene transferase activity of this metalloprotein is severely inhibited by it, imposing the need for strictly anaerobic conditions to conduct these reactions. In this report, we describe how substitution of the native heme cofactor with an iron-chlorin e6 complex enabled the development of a biocatalyst capable of promoting the cyclopropanation of vinylarenes with high catalytic efficiency (up to 6970 TON), turnover rate (>2000 turnovers/min), and stereoselectivity (up to 99% de and ee) in the presence of oxygen. The artificial metalloenzyme can be recombinantly expressed in bacterial cells, enabling its application also in the context of whole-cell biotransformations. This work makes available a robust and easy-to-use oxygen-tolerant biocatalyst for asymmetric cyclopropanations and demonstrates the value of porphyrin ligand substitution as a strategy for tuning and enhancing the catalytic properties of hemoproteins in the context of abiological reactions

Chemoselective Cyclopropanation over Carbene Y–H Insertion Catalyzed by an Engineered Carbene Transferase

Hemoproteins have recently emerged as promising biocatalysts for promoting a variety of carbene transfer reactions including cyclopropanation and Y–H insertion (Y = N, S, Si, B). For these and synthetic carbene transfer catalysts alike, achieving high chemoselectivity toward cyclopropanation in olefin substrates bearing unprotected Y–H groups has proven remarkably challenging due to competition from the more facile carbene Y–H insertion reaction. In this report, we describe the development of a novel artificial metalloenzyme based on an engineered myoglobin incorporating a serine-ligated Co-porphyrin cofactor that is capable of offering high selectivity toward olefin cyclopropanation over N–H and Si–H insertion. Intramolecular competition experiments revealed a distinct and dramatically altered chemoselectivity of the Mb­(H64V,V68A,H93S)­[Co­(ppIX)] variant in carbene transfer reactions compared to myoglobin-based variants containing the native histidine-ligated heme cofactor or other metal/proximal ligand substitutions. These studies highlight the functional plasticity of myoglobin as a “carbene transferase” and illustrate how modulation of the cofactor environment within this metalloprotein scaffold represents a valuable strategy for accessing carbene transfer reactivity not exhibited by naturally occurring hemoproteins or transition metal catalysts

Cyclopropanations via Heme Carbenes: Basic Mechanism and Effects of Carbene Substituent, Protein Axial Ligand, and Porphyrin Substitution

Catalytic carbene transfer to olefins is a useful approach to synthesize cyclopropanes, which are key structural motifs in many drugs and biologically active natural products. While catalytic methods for olefin cyclopropanation have largely relied on rare transition-metal-based catalysts, recent studies have demonstrated the promise and synthetic value of iron-based heme-containing proteins for promoting these reactions with excellent catalytic activity and selectivity. Despite this progress, the mechanism of iron-porphyrin and hemoprotein-catalyzed olefin cyclopropanation has remained largely unknown. Using a combination of quantum chemical calculations and experimental mechanistic analyses, the present study shows for the first time that the increasingly useful CC functionalizations mediated by heme carbenes feature an Fe^II-based, nonradical, concerted nonsynchronous mechanism, with early transition state character. This mechanism differs from the Fe^IV-based, radical, stepwise mechanism of heme-dependent monooxygenases. Furthermore, the effects of the carbene substituent, metal coordinating axial ligand, and porphyrin substituent on the reactivity of the heme carbenes was systematically investigated, providing a basis for explaining experimental reactivity results and defining strategies for future catalyst development. Our results especially suggest the potential value of electron-deficient porphyrin ligands for increasing the electrophilicity and thus the reactivity of the heme carbene. Metal-free reactions were also studied to reveal temperature and carbene substituent effects on catalytic vs noncatalytic reactions. This study sheds new light into the mechanism of iron-porphyrin and hemoprotein-catalyzed cyclopropanation reactions and it is expected to facilitate future efforts toward sustainable carbene transfer catalysis using these systems